Advances in gene recombinant technology have enabled the provision of a stable supply of various protein formulations. In particular, a variety of recombinant antibody drugs, which are more selective than normal-drugs, have been developed and entered clinical trial in recent years.
In these recombinantly produced formulations containing physiologically active proteins, there is a need to remove host DNA and contaminant DNA associated with viral contamination. Under present World Health Organization (WHO) criteria, the amount of DNA in biological drugs should not exceed 100 pg DNA/dose. To meet this criteria, in general, an aqueous medium containing a physiologically active protein obtained from host cells is treated by anion-exchange chromatography, hydroxyapatite chromatography or a combination thereof for the purpose of removing DNA.
In particular, in a case where a physiologically active protein is an antibody produced recombinantly in mammalian host cells, the aqueous medium is treated by affinity column chromatography on Protein A or Protein G before purification by various types of chromatography, based on the binding property of Protein A or Protein G to IgG Fc chain.
By way of example, in JP 5-504579 A, an antibody-containing aqueous medium obtained from mammalian cell culture is applied to Protein A/G column chromatography to adsorb antibody molecules onto the column, and is then eluted with an acidic solution (about 0.1 M citric acid, pH 3.0-3.5) to release the antibody molecules. The resulting acidic eluate is sequentially applied to ion-exchange column chromatography and size exclusion column chromatography to give the purified antibody molecules.
However, these individual chromatographic processes and a combination thereof are time-, labor- and cost-consuming, as well as being complicated. Moreover, they fail to provide stable results.
Thus, there is a need to develop a simpler and less expensive method for purifying physiologically active proteins, especially antibodies, which can ensure removal of contaminant DNA, and which can minimize a loss of physiologically active proteins.